The proposed work examines the possibility that a soluble Ca 2 ion dependent regulatory protein (CDR) participates in the biochemical events initiated by the binding of opiates to membrane receptors, leading to analgesia and the tolerant dependent state. The CDR, which is present in high concentration in the brain, regulates both phosphodiesterase and adenylate cyclase activities. The association of the CDR with neural membranes is a dynamic process dependent on Ca2 ion and the phosphorylation of the membranes. Since opiates affect Ca2 ion levels, adenylate cyclase activity and phosphorylation of membranes, it is likely that the CDR is involved in the action of opiates. A study of the effects of acute and chronic opiate administration on the levels of CDR in the brains of rats and the distribution of the protein between membranes and cytoplasm may provide support for this hypothesis. In order to gain additional insight, the binding of 125I-CDR to membranes and the direct binding of opiates to the CDR in vitro, will be examined. Copper is a potent inhibitor of opiate receptor binding and Cu2 ion chelators potentiate the analgesic effects of morphine in vovo. We therefore plan to investigate the effects of opiates on specific Cu2 ion binding sites of membranes using the ability of Cu2 ion to quench native membrane fluorescence as a measure of the relative amount of bound Cu2 ion.